Baculovirus expression and immunological detection of the major structural proteins of porcine reproductive and respiratory syndrome virus.
Each of the three major structural proteins (envelope glycoprotein E, nonglycosylated membrane protein M, and nucleoprotein N) of an American strain of porcine reproductive and respiratory syndrome virus (PRRSV) was expressed using a recombinant baculovirus expression system. Insect cells infected with the respective recombinant baculovirus synthesized five distinct forms of glycoprotein E with a molecular mass (M(r)) of either 17, 20, 23, 25 or 26 K, and a single form of nonglycosylated protein M and nucleocapsid N with a M(r) of approximately 21 and 15 K, respectively. Because the number of forms of the glycoprotein E was reduced from five to two (20 and 17 K) when infected cells were treated with tunicamycin, we speculate that the 23, 25 and 26 K forms represent different degrees of glycosylation of the same protein, and that the 20 and 17 K peptides represent nonglycosylated forms with and without, respectively, the N-terminal signal sequence. All the proteins were identified by immunoblot with convalescent sera from animals infected with an American strain of PRRSV, indicating that they were similar to the native proteins. The recombinant proteins were purified and used to induce monospecific antisera in rabbits. The ability to produce each protein in the baculovirus system provides an additional means for their structural and functional characterization.
Kreutz
L C
LC
Virology Swine Research Unit, National Animal Disease Center, USDA, Agricultural Research Service, Ames, IA 50010, USA. Lckreutz@creta.ccr.ufsm.br
Mengeling
W L
WL
D000818:Animals; D000906:Antibodies; D016367:Baculoviridae; D015153:Blotting, Western; D002460:Cell Line; D003001:Cloning, Molecular; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D019084:Fluorescent Antibody Technique, Indirect; D008970:Molecular Weight; D016133:Polymerase Chain Reaction; D019316:Porcine respiratory and reproductive syndrome virus / Q000235:genetics / Q000378:metabolism*; D011817:Rabbits; D011994:Recombinant Proteins / Q000032:analysis / Q000096:biosynthesis; D018411:Spodoptera; D013552:Swine; D014162:Transfection; D015678:Viral Structural Proteins / Q000032:analysis / Q000096:biosynthesis*
D000906:Antibodies; D017931:DNA Primers; D011994:Recombinant Proteins; D015678:Viral Structural Proteins